This rat study (n=180) investigated the pharmacokinetic interactions between 5-MeO-DMT (0.25mg/0.25kg) and the MAO-A inhibitor clorgyline and the MAO-A/B inhibitor pargyline, and their effects on prepulse inhibition (PPI), a phenomenon related to the ability to filter out unnecessary information. Results confirmed that the MAO inhibitors increase the accumulation of 5-MeO-DMT in the nervous system, and boost its disruptive effects on PPI by activating the 5-HT_2a receptor pathway.

Abstract

“Introduction: Monoamine oxidase inhibitors (MAOIs) are often ingested together with tryptamine hallucinogens, but relatively little is known about the consequences of their combined use. We have shown previously that monoamine oxidase-A (MAO-A) inhibitors alter the locomotor profile of the hallucinogen 5-methoxy-N,N-dimethyltryptamine (5-MeO-DMT) in rats, and enhance its interaction with 5-HT_2A receptors. The goal of the present studies was to investigate the mechanism for the interaction between 5-MeO-DMT and MAOIs, and to determine whether other behavioral responses to 5-MeO-DMT are similarly affected. Hallucinogens disrupt prepulse inhibition (PPI) in rats, an effect typically mediated by 5-HT_2A activation. 5-MeO-DMT also disrupts PPI but the effect is primarily attributable to 5-HT_1A activation.

Methods: The present studies examined whether an MAOI can alter the respective contributions of 5-HT_1A and 5-HT_2A receptors to the effects of 5-MeO-DMT on PPI. A series of interaction studies using the 5-HT_1A antagonist WAY-100635 and the 5-HT_2A antagonist MDL 11,939 were performed to assess the respective contributions of these receptors to the behavioral effects of 5-MeO-DMT in rats pretreated with an MAOI. The effects of MAO-A inhibition on the pharmacokinetics of 5-MeO-DMT and its metabolism to bufotenine were assessed using liquid chromatography–electrospray ionization–selective reaction monitoring–tandem mass spectrometry (LC-ESI-SRM-MS/MS).

Results: 5-MeO-DMT (1 mg/kg) had no effect on PPI when tested 45-min post-injection but disrupted PPI in animals pretreated with the MAO-A inhibitor clorgyline or the MAO-A/B inhibitor pargyline. The combined effect of 5-MeO-DMT and pargyline on PPI was antagonized by pretreatment with either WAY-100635 or MDL 11,939. Inhibition of MAO-A increased the level of 5-MeO-DMT in plasma and whole brain, but had no effect on the conversion of 5-MeO-DMT to bufotenine, which was found to be negligible.

Discussion: The present results confirm that 5-MeO-DMT can disrupt PPI by activating 5-HT_2A, and indicate that MAOIs alter 5-MeO-DMT pharmacodynamics by increasing its accumulation in the central nervous system.”

Author: Adam L. Halberstadt

Summary

Introduction

Tryptamine hallucinogens such as DMT and 5-MeO-DMT are commonly abused together with drugs that inhibit monoamine oxidase-A (MAOA). This is because tryptamines are normally inactive orally due to first-pass metabolism, but -carbolines contribute to the activity of ayahuasca by blocking the catabolism of DMT in the gastrointestional tract.

Hallucinogens bind to 5-HT2A receptors, but with differing selectivity. Phenylalkylamines are selective for 5-HT2 sites, whereas indoleamines are non-selective for 5-HT2A receptors.

When administered to rats pretreated with an MAOI, 5-MeO-DMT produces biphasic effects, initially suppressing locomotor activity and then increasing activity at later time points. This effect can be blocked by the selective 5-HT2A antagonist MDL 11,939 but not by WAY-100,635.

MAOA inhibition alters the pharmacokinetics of 5-MeO-DMT, which may explain why MAOA inhibitors increase blood and brain concentrations of 5-MeO-DMT. Nevertheless, MAOA inhibition does not directly mediate the delayed hyperactivity of 5-MeO-DMT.

The behavioral profile of 5-MeO-DMT is altered in the presence of an MAOI, because 5-MeO-DMT is deaminated by MAOA and O-demethylated to bufotenine by cytochrome P450 2D6. Bufotenine is a potent 5-HT2A agonist.

The present studies assessed the interaction between 5-MeO-DMT and MAOIs, and determined whether other behavioral responses to 5-MeO-DMT are similarly affected. They also assessed whether inhibition of MAOA alters the pharmacokinetics of 5-MeO-DMT and its biotransformation to bufotenine.

2.1. Animals

Male Sprague – Dawley rats were housed in pairs in a temperature- and humidity-controlled vivarium under a 12-h reverse light – dark cycle. They were provided food and water ad libitum.

Serotonin was measured using NMS, clorgyline, pargyline HCl, bufotenine, -Phenylethyl-4-piperidinemethanol, methanol, and ethyl acetate. Blank rat plasma was purchased from Biochemed.

For the in vivo experiments, drugs were administered subcutaneously in a volume of 1 mL/kg. 5-MeO-DMT, WAY-100,635, clorgyline, and pargyline were dissolved in isotonic saline.

2.3.1. Behavioral pattern monitor

Rats were placed in a black Plexiglas chamber with 2.5 cm holes in the walls and floor. Infrared photobeams were used to detect nosepokes and rearings were detected by touchplates on the walls.

2.3.2. Acoustic startle

Startle chambers were used to measure startle reactivity. A high-frequency loudspeaker was mounted 24 cm above the Plexiglas stabilimeter, and 100 1-ms samples were recorded at the onset of the startling stimulus.

Acoustic startle test sessions consisted of startle trials and prepulse trials. Animals were acclimated to 65 dB broadband noise for 5 min, and then tested in a startle/PPI session for 1 week to create treatment groups matched for baseline levels of startle and PPI.

2.4.1. Acoustic startle

In experiments 1 and 2, rats were treated with pargyline, clorgyline, or WAY-100,635 (20 min) before 5-MeO-DMT (0, 0.1, or 1.0 mg/kg), and in experiments 3 and 4, rats were treated with MDL 11,939 (20 min) before 5-MeO-DMT (0, 1.0 mg/kg)

2.4.2. Behavioral pattern monitor

Rats were pretreated with vehicle or clorgyline 20 min before administration of 5-MeO-DMT, and locomotor activity was monitored for 10, 20, 30, 40, 50, or 60 min. Brains were removed, flash frozen in isopentane at 80 °C, and stored at 40 °C.

2.5.1. Preparation of calibrators and controls

The calibrators and controls were prepared in silanized, 16 100 mm glass culture tubes containing 1 mL blank rat plasma. Separate sets of working solutions were used for the calibrators and controls.

2.5.2. Preparation of samples for analysis

For the plasma samples, 1 mL of each sample was transferred to a silanized 16 100 mm glass culture tube, and 10 fold diluted brain tissue homogenates were prepared for each brain. The homogenates were thawed, and 1 mL of each sample homogenate was transferred to a new 16 100 mm glass culture tube.

2.5.3. Extraction

Clean Screen ZSDAU020 solid phase extraction columns were used for the extraction procedure. The extracts were reconstituted with 100 l of 10 mM ammonium acetate (pH 5.0)/methanol (80:20) and then transferred to separate 300 l conical, polypropylene autosampler vials with snap caps.

2.5.4. LC–MS/MS analysis

A liquid chromatography – tandem mass spectrometry (LC – MS/ MS) system was used to analyze 5-MeO-DMT and bufotenine in extracts prepared from the study samples. A calibration curve was generated from the analysis of the calibration standards and LCquan 2.5 data analysis software was used for the quantification.

2.6.1. Acoustic startle

The amount of PPI was calculated for each PREPULSE + PULSE trial type, and the level of spontaneous motor activity was assessed using NOSTIM trials. ANOVA was used to analyze data, and significance was demonstrated by surpassing an alpha level of 0.05.

2.6.2. Behavioral pattern monitor

Locomotor activity was quantified by crossings between eight equal square sectors within the BPM. Data were analyzed using two-way ANOVA with clorgyline pretreatment and time block as between-subject factors.

5-MeO-DMT and bufotenine concentrations were measured in rats. The area under the curve (AUC) was calculated for each group and compared using Kaplan – Meier survival analysis and the Tarone – Ware test.

5-MeO-DMT was shown to disrupt PPI when administered 10 min prior to startle testing. It was also shown that 5-MeO-DMT altered PPI when administered 45 min prior to testing, and that the response to 5-MeO-DMT was altered in animals pretreated with 10 mg/kg pargyline.

Pretreatment with pargyline significantly reduced the amplitude of the startle response, but treatment with 5-MeO-DMT had no effect on the startle response. The combination of pargyline and 5-MeO-DMT reduced PPI in subgroups matched for startle level.

3.2. Effect of clorgyline and 5-MeO-DMT on acoustic startle and sensorimotor gating

We tested whether the response to 5-MeO-DMT is altered in animals pretreated with 0.3 mg/kg clorgyline. 5-MeO-DMT significantly reduced PPI in animals pretreated with clorgyline at all three prepulse intensities.

3.3. Involvement of 5-HT1A and 5-HT2A receptors in the effects of pargyline and 5-MeO-DMT on sensorimotor gating

The effects of pargyline and 5-MeO-DMT on PPI were determined by blocking 5-HT1A and 5-HT2A receptors, and the combination of pargyline and 5-MeO-DMT significantly reduced PPI at all three prepulse intensities in vehicle-pretreated animals.

3.3.1. WAY-100,635

For PPI, pretreatment with 1 mg/kg WAY-100,635 antagonized the reduction produced by 10 mg/kg pargyline and 1 mg/kg 5-MeO-DMT, but the reduction produced by 1 mg/kg WAY-100,635 did not achieve significance.

3.3.2. MDL 11,939

Pargyline and 5-MeO-DMT significantly reduced PPI when pretreated with MDL 11,939, but did not alter startle amplitude.

3.4.1. Analysis method

Several different extraction procedures were investigated, but were associated with poor recovery and sensitivity. We used liquid chromatography – electrospray ionization – selective reaction monitoring – tandem mass spectrometry for the simultaneous detection of 5-MeO-DMT and bufotenine in plasma and brain. Chromatography of 5-MeO-DMT, bufotenine, and NMS was subject to ion suppression, but retention times were not affected, and there was little variation within or between runs. Calibration was reproducible, and concentrations were within 5% of target with %CVs within 12.9% for both analytes.

Rats were administered 1 mg/kg 5-MeO-DMT after pretreatment with vehicle or clorgyline. The concentration of 5-MeO-DMT in plasma and brain peaked within the first 20 min after SC administration and then rapidly declined. Clorgyline significantly increased the Cmax of 5-MeO-DMT in whole brain, and 5-MeO-DMT was also detectable for a longer period of time in the presence of an MAO inhibitor. Bufotenine concentrations in plasma and brain were not altered by clorgyline pretreatment.

To confirm the expected behavioral interaction between 0.3 mg/kg clorgyline and 1 mg/kg 5-MeO-DMT, locomotor activity was assessed in the BPM prior to collection of analytical samples. Clorgyline pretreatment significantly increased locomotor activity.

Discussion

Pretreatment with an MAOI markedly prolongs the behavioral response to 5-MeO-DMT. 5-MeO-DMT disrupts PPI in rats by activating both 5-HT1A and 5-HT2A receptors, and this effect is dependent on activation of both 5-HT1A and 5-HT2A receptors.

Pretreatment with the MAOA inhibitor clorgyline dramatically altered the pharmacokinetics of 5-MeO-DMT in rats. This was probably because clorgyline blocked the metabolism of 5-MeO-DMT and increased the concentration of 5-MeO-DMT in blood and brain 40-fold and almost 100-fold, respectively.

5-MeO-DMT produces brief alterations of locomotor activity and PPI in rats, but its effects are prolonged and involve 5-HT2A receptors. Yu and colleagues showed that MAOIs increase the O-demethylation of 5-MeO-DMT in mice, resulting in elevated and prolonged systemic exposure to bufotenine. The present results reveal that MAOIs alter the behavioral response to 5-MeO-DMT by blocking its deamination.

The delayed 5-HT2A-mediated behavioral response induced by 5-MeO-DMT and an MAOI may be explained by the delayed sensitization of dopaminergic transmission, which would explain why 5-HT2A antagonists block the behavioral response at 45 min post-treatment, but not at earlier time-points.

The present findings confirm that 5-MeO-DMT can activate 5-HT2A receptors in vivo. It has also been shown that 5-MeO-DMT can induce delayed hyperactivity via 5-HT2A receptor activation.

The 5-HT2 agonist 5-MeO-DMT activates 5-HT2A receptors in rats in vivo, but the 5-HT1A receptor inhibits 5-HT2A-mediated behavioral responses. However, in the presence of an MAO inhibitor 5-MeO-DMT accumulates in the brain at levels that are adequate to increase locomotor activity and disrupt PPI via 5-HT2A.

5-MeO-DMT can induce symptoms of serotonin syndrome in rats, but the duration of the serotonin syndrome is shorter than the interval between drug treatment and startle testing, demonstrating that the serotonin syndrome did not interfere with expression of the startle response.

MAOIs can alter the psychoactive effects of 5-MeO-DMT and other tryptamines, and polypharmacy has been used to experiment with the effects of these drugs. 5-MeO-DMT inhibits 5-HT uptake at low micromolar concentrations, and this effect could contribute to interactions with MAOA inhibitors and other 5-HT agonists. The high concentrations of 5-MeO-DMT present in the brain may also interact with a variety of other targets, potentially producing adverse effects and toxicity.

Acknowledgments

This work was supported by grants from NIMH, NIDA, and the Brain and Behavior Research Foundation.